Research: Transcriptional regulatory proteins function as repressors or activators of gene expression and play a paramount role in cellular differentiation. The overall goal of this proposal is to increase our understanding of transcriptional regulatory mechanisms that govern differentiation of cardiac and skeletal myocytes. The winged helix or forkhead/HNF-3 gene family is comprised of several recently identified proteins that function in pattern formation in the early embryo. Preliminary studies in this laboratory have identified a novel member of the winged helix family that is expressed in cardiac and skeletal muscle, termed myocyte nuclear factor (MNF). MNF exhibits sequence-specific DNA binding to a positive transcriptional control element within a muscle- specific enhancer region from the human myoglobin gene. The major hypothesis to be addressed by this grant proposal is that MNF and/or closely related winged helix proteins play a significant role in differentiation of skeletal and cardiac myocytes. The following four specific aims are proposed: l) to clone the mouse genomic locus encoding MNF; 2) to identify and to define the relationships between members of the winged helix gene family that are expressed in cardiac and skeletal muscle (i.e. variants generated by alternative splicing); 3) to examine the spatial and temporal distribution of MNF during embryogenesis; 4) to determine the functional importance of winged helix proteins during development by gene disruption using homologous recombination in embryonic stem cells and subsequent generation of strains of mice bearing homozygous null mutations in the gene(s) encoding these proteins. Achievement of these aims will define the role of MNF in cardiac and skeletal myogenesis. In the long term, advances in our understanding of the control of muscle development may foster novel strategies to improve cardiac function in patients with cardiovascular disease.